Synaptic transmission is a critical feature of the nervous system and is as such tightly regulated on many different time-scales. The ubiquitin-proteasome system (UPS) plays an important role in synaptic transmission by maintaining appropriate protein homeostasis. The UPS regulates synapse formation during development, by shaping long-term plasticity, by altering the number of active synapses and by changing the size of vesicle pools. All of these regulatory functions occur on a time scale of hours to days and are consistent with the central role of the UPS in protein degradation: covalent attachment of ubiquitin tags proteins for degradation by the proteasome. We have recently discovered that the UPS also regulates neurotransmitter release on a time-scale of a few minutes or less. This led us to the hypothesis that ubiquitination at the synapse rapidly regulates release by changing the activity level of target proteins rather than changing their half-life. Thus, protein ubiquitination can serve as an acute dynamic regulator of synaptic function. This represents a novel regulatory pathway that is of clear basic science interest given the central role of synaptic transmission in nervous system function. However, the pathway is also of clear clinical interest as it might offer insight into the earliest steps of dysfunction associated with neurodegenerative diseases. In many neurodegenerative diseases signs of synaptic dysfunction precede clinical symptoms. Neurodegenerative diseases are also characterized by a UPS that is impaired either through mutations in the pathway, by misfolded proteins or via environmental toxins. The recognition of the rapid regulatory role of the UPS in shaping synaptic transmission might offer an early causal link between UPS impairment and synaptic dysfunction. In this grant application we will test the physiological mechanisms that mediate the rapid regulation of presynaptic release by ubiquitination (Aim 1). Together with the identification of the synaptic ubiquitome (Aim2) these experiments will enable discovery of pathways and processes that are of basic science interest and that might provide novel druggable targets for neurological diseases.